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Abstract

Sprouty1 was identified from a genomic screen of human heart failure samples for novel regulators of receptor tyrosine kinase signaling. In silico Sprouty1 promoter analysis revealed three putative hypoxia response elements potentially capable of binding hypoxia inducible factor-1α (HIF-1α). The first goal of this study was to test the hypothesis that Sprouty1 expression was increased under hypoxic conditions through HIF-1α. Using HL-1 cardiac myocytes, we identified an increase in Sprouty1 mRNA expression after exposure to 1% oxygen for 12 hours (n = 6, p < 0.01). The observed increase in Sprouty1 transcript was attenuated by a dominant negative HIF-1α mutant with respect to control transfected cells (n = 6, p < 0.01). Furthermore, we also observed a corresponding increase in Sprouty1 protein levels at 12 hours under the same oxygen-limiting conditions. To understand the functional relevance of hypoxia-mediated Sprouty1 up-regulation on cell survival, neonatal cardiac myocytes were infected with Sprouty1 adenovirus. The percentage of apoptotic nuclei under hypoxic conditions was three-fold greater in Sprouty1 infected cardiac myocytes compared to control (n = 3, p < 0.05). Interestingly, an association was observed between the expression of Sprouty1 and the pro-apoptotic gene BNIP3, itself induced in hypoxic cardiac myocytes. Adenoviral up-regulation of Sprouty1 alone, in the absence of hypoxia, was sufficient to stimulate BNIP3 expression. In summary, this data suggests that hypoxia stimulates expression of Sprouty1 in cardiac myocytes and potentiates apoptosis in association with increased expression of the pro-apoptotic gene, BNIP3. To our knowledge, this is the first report that BNIP3 expression is increased by Sprouty1, a conserved inhibitor of receptor tyrosine kinase signaling. Therefore, these findings may have important implications in the cross-talk and integration between receptor tyrosine kinase signaling, apoptotic regulatory genes, and cell fate in the setting of hypoxia/ischemia.